This project will demonstrate a powerful new strategy for diagnosing infectious diseases in minimally equipped laboratories or at point of care (POC). Developed for biodefense and similar low-complexity testing purposes, the RAMP(R) antigen detection system (Response Biomedical Corporation, Vancouver, BC, Canada) will be adapted for POC diagnosis of tuberculosis (TB). The RAMP system has been extensively evaluated by independent testers, and its high sensitivity and outstanding robustness have been demonstrated. Its sensitivity will be enhanced by integration with a novel electrohydrodynamic microtip system designed to selectively concentrate Mycobacterium tuberculosis (MTB) cells in sputum samples prior to RAMP analysis. The microtip system was developed and validated by NanoFacture, Inc., in collaboration with the Nanomanufacturing Laboratory at the University of Washington. The integrated microtip-RAMP test will detect MTB cells with a lower limit of detection of <100 cfu per mL of sputum, comparable to PCR in a 20 min total test time. Specific aims of the project are: 1) To develop a RAMP test for MTB cells in sputum;2) to develop sample processing procedures that utilize the microtip system to concentrate MTB cells from human sputum;and 3) to evaluate the performance of the microtip-RAMP system in a field trial in Dhaka, Bangladesh. Targeted pre-clinical performance for the 20-min microtip-RAMP test will have sensitivity and specificity of >90% relative to culture. In addition, the hypothesis will be tested that microtip-RAMP is significantly more sensitive than AFB smear microscopy, currently the world's most widely used rapid test to detect MTB cells in sputum.